We propose a genetic analysis of three herpes simplex virus functions (i) the role of Beta polypeptide ICP6, (ii) HSV-2 morphological transformation, and (iii) the locus synl-2. We will create mutants in cloned DNA fragments by using a variety of in vitro mutagenic techniques. We will study the viral genes by analysis of mutants either in in vitro transfection-transformation in vivo after transfer of constructed lesions into viral genomes. The function of ICP6 will be studied in experiments designed to determine the consequences of lesions in this potential Beta regulatory gene. We hypothesize that ICP6 expression may be an important regulator of Alpha gene expression. Thus, the proposed protein synthetic experiments with ICP6 mutants will examine ICP6 experssion in the viral lytic cycle and the effects of its expression on Alpha polypeptide synthesis. HSV-2 morphological transformation will be studied by examining the ability of a series of mutated Bg1II-N cloned DNA fragment to transform primary or established rodent cell lines in in vitro transfection-transformation assays. The selection for transformed cells will be aided by linked transfer with Ecogpt sequences (a fragment containing the template for E. coli xanthine-guanine phosphoribosyl-transferase [XGPRT]) and preselection for XGPRT expression. The gene that maps at the exact position of the transforming sequences will be determined by two approaches (i) analysis of polypeptides specified by intertypic recombinant in the region of the transforming function, and (ii) by in vitro translation of selected transcripts which are homologous to the exact sequences of the morphological transformation region. This information is essential to the understanding of HSV-2 transformation. The function and identification of the product of the HSV locus synl-2 (previously synl, syn2) will be studied in experiments to determine whether any template other than alpha polypeptide ICP27 maps at that position. We propose to create additional mutations in the locus synl02 to study the interaction of ICP27 on a functional level with known structural features of the HSV genome at the position of the comapping ICP27 template and locus synl-2.